Table of Contents

Volume 5, Issue 8

  April 15, 2006

  Pages 797 - 901

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 Issue Catalogue (MARC XML)

EXTRA VIEWS

Open Access Article

CARD Tricks: Controlling the Interactions of CARD6 with RICK and Microtubules

Almut Dufner and Tak W. Mak
Pages 797 - 800
http://dx.doi.org/10.4161/cc.5.8.2635
Abstract | PDF

In recent years, a number of proteins have been identified that contain a homotypic interaction motif called the caspase recruitment domain (CARD). Most proteins containing a CARD are involved in pathways regulating apoptosis or adaptive or innate immunity. Examples of prominent CARD proteins are caspase-9 and Apaf1, which are involved in the intrinsic death pathway; BCL10 and CARD11, which mediate antigen receptor- induced NF-?B activation; and receptor-interacting protein (RIP)-like interacting caspase-like apoptosis regulatory protein kinase (RICK) and the nucleotide-binding oligomerization domain (NOD) proteins, which induce NF-?B activation in response to intracellular bacterial peptidoglycan. The most recently discovered pathway involving CARD proteins senses virally-derived double-stranded (ds) RNA and initiates a host defense signaling program. CARD6 is a CARD-containing protein with a domain structure not shared by any other CARD protein. Although the CARD6 cDNA was deposited in GenBank five years ago, the physiological function of full-length CARD6 has yet to be reported. Here we review our initial characterization of CARD6 and discuss the functional implications of various conserved modules found in the CARD6 protein sequence. We conclude that CARD6 is structurally and potentially functionally related to the superfamily of interferon (IFN)-inducible GTPases, a growing family of host defense proteins that confer cell-autonomous immunity.

Open Access Article

Oncogenes and the DNA Damage Response: Myc and E2F1 Engage the ATM Signaling Pathway to Activate p53 and Induce Apoptosis

Sungki Hong, Raju V. Pusapati, John T. Powers and David G. Johnson
Pages 801 - 803
http://dx.doi.org/10.4161/cc.5.8.2638
Abstract | PDF

Activation of the ATM DNA damage response pathway is commonly observed in a variety of early-stage neoplasias. It has been proposed that this checkpoint response functions to suppress the development of cancer. A recent report from our laboratory demonstrates that ATM does indeed function to suppress tumorigenesis by responding to at least some oncogenic stresses. Transgenic expression of Myc is found to cause DNA damage in vivo and ATM is shown to respond to this damage by inducing the accumulation and phosphorylation of p53. In the absence of ATM, p53- dependent apoptosis is reduced and epithelial tumorigenesis is accelerated in Myc transgenic mice. Deregulated expression of the E2F1 transcription factor also elicits an ATM-dependent checkpoint response that activates p53 and promotes apoptosis, although the mechanism by which E2F1 and Myc stimulate ATM may differ. These findings have relevance for understanding why the ATM pathway is activated in many human cancers, what generates the selective pressure for p53 inactivation during tumorigenesis, and why AT patients and carriers are predisposed to developing cancer.




PERSPECTIVES

Open Access Article

TRAF3: A New Regulator of Type I Interferons

Supriya K. Saha and Genhong Cheng
Pages 804 - 807
http://dx.doi.org/10.4161/cc.5.8.2637
Abstract | PDF

Type I interferons are a major and essential component of the mammalian antiviral response. While many cell types produce type I interferons following viral infection, how cells detect the virus has remained a mystery for many years. Recently, multiple genome-encoded viral recognition receptors have been identified. Interestingly, all of the major viral recognition pathways characterized thus far require TRAF3 to initiate type I interferon production. In this review, we comment on the mechanistic and biological implications for the new role of TRAF3 in innate antiviral immunity.

Open Access Article

Clonal Expansion in the Human Gut: Mitochondrial DNA Mutations Show Us the Way

Stuart A.C. McDonald, Sean L. Preston, Laura C. Greaves, Simon J. Leedham, Matthew A. Lovell, Janusz A.Z Jankowski, Douglass M. Turnbull and Nicholas A. Wright
Pages 808 - 811
http://dx.doi.org/10.4161/cc.5.8.2641
Abstract | PDF

The mechanisms of how DNA mutations are fixed within the human gastrointestinal tract and how they spread are poorly understood and are hotly debated. It has been well documented that human colonic crypts are clonal units; one stem cell within the crypt becoming dominant and taking over the crypts’ entire stem cell population – so called monoclonal conversion. Studies have revealed that crypts can exist as families and develop into patches. The questions have been how do such patches in the human colon develop? Does this have implications on how DNA mutations spread? We have previously shown that mitochondrial DNA (mtDNA) mutations, which result in the deficiency of cytochrome c oxidase, are established within a single colonic crypt stem cell, resulting in a crypt with a mixed phenotype. Over time that mutated stem cell can take over the entire stem cell population resulting in a wholly-mutated crypt. We have furthered this research by showing that entirely cytochrome c oxidase–deficient crypts are able to divide by a process called crypt fission, to form two cytochrome c oxidasedeficient daughter crypts, each sharing the exact parental mtDNA mutation. Furthermore, patches of these crypts also possess a founder mtDNA mutation suggesting that fission repeats itself to form patches, which increase in size with age. Here, we hypothesize that this can be expanded into other areas of the gastrointestinal tract, especially the stomach, where there is a paucity of data regarding clonality and the spread of DNA mutations. We ask if these mutated crypts expand at a different rate to wild type ones. We also discuss the implications for the spread of potential carcinogenic mutations within the gut.

Open Access Article

Lymphatic or Hematogenous Dissemination: How Does a Metastatic Tumor Cell Decide?

Sunny Y. Wong and Richard O. Hynes
Pages 812 - 817
http://dx.doi.org/10.4161/cc.5.8.2646
Abstract | PDF

The formation of distant metastases is the deadliest phase of cancer progression. Although numerous studies have identified genes and mechanisms that affect metastasis after tumors have reached secondary sites, our knowledge about how cancer cells initially gain access to systemic circulation is limited. Since tumors can enter the blood directly by intravasating into venous capillaries or indirectly via lymphatics, it is important to evaluate the relative contributions of both pathways as routes of egress from the primary site. Insights into tumor and stromal factors governing the intravasation process may help explain why certain tumors exhibit “preferred” pathways for metastatic dissemination, both clinically and in experimental animal models.

Open Access Article

Chromosomal Instability, Colorectal Cancer and Taxane Resistance

Charles Swanton, Ian Tomlinson and Julian Downward
Pages 818 - 823
http://dx.doi.org/10.4161/cc.5.8.2682
Abstract | PDF

mitosis. Aberrations in key regulators of this process have been found in human malignancies associated with chromosomal instability (CIN). Important chemotherapeutic agents which alter microtubule dynamics such as paclitaxel and vincristine, prolong spindle checkpoint activation and delay the completion of mitosis. Intriguingly, inhibition of BubR1 or overexpression of Aurora kinase A, spindle checkpoint regulators implicated in CIN, promote resistance to microtubule inhibitors (MTIs) in vitro. Taxanes have failed to demonstrate significant clinical benefit in phase II trials in colorectal cancer (CRC). The high incidence of CIN in this disease, coupled with alterations in spindle checkpoint regulators in vivo, may explain the disappointing results associated with taxane based therapies for CRC. A phase II trial of taxanes in patients with metastatic CIN negative CRC may be indicated.

Open Access Article

p38 MAPK in Development and Cancer

Cynthia Bradham and David R. McClay
Pages 824 - 828
http://dx.doi.org/10.4161/cc.5.8.2685
Abstract | PDF

p38 is a MAPK that has been shown to induce a wide variety of biological effects in cell culture, in response to a wide range of stimuli. These effects are dependent not only on the stimuli, but also on the cellular context, resulting in a bewildering array of possibilites1. For example, p38 was shown to induce apoptosis in some cells, but prevent apoptosis in others2. Similarly opposed effects had been observed with respect to cell cycle regulation2. The role of p38 in inflammatory disease has been appreciated from the beginning, since it was initially identified as an cytokine inducer3. More recently, p38 function has been evaluated in vivo, and through these studies p38 has emerged as an important regulator of both embryonic development and cancer progression. This review will focus on these in vivo studies in an effort to provide perspective on p38 biologically and as a pharmacological target.

Open Access Article

p8 and Prothymosin Alpha : Unity is Strength

Cédric Malicet, Jean Charles Dagorn, José Luis Neira and Juan L. Iovanna
Pages 829 - 830
http://dx.doi.org/10.4161/cc.5.8.2686
Abstract | PDF

p8 and prothymosine a are two natively unstructured proteins with anti-apoptotic activity. We showed that their interaction results in the formation of a one-to-one heterodimer complex with stable structure. To test whether the heterodimer bears the function previously attributed to both proteins, we monitored the consequences on apoptosis of modulating in vitro the concentrations of both proteins. Over-expression was obtained by transfection of appropriate vectors and inhibition by using specific siRNAs. In all conditions inhibition of apoptosis correlated with the level of the partner with lowest concentration, demonstrating that the anti-apoptotic effect previously attributed to each proteins was in fact borne by the p8/ProTa complex, the two proteins, being individually inactive. These results show that the function attributed to a natively unfolded protein might actually belong to a multi-protein complex in which the protein of interest is engaged.

Open Access Article

Check, Double Check: The G2 Barrier to Cancer

Floris Foijer and Hein te Riele
Pages 831 - 836
http://dx.doi.org/10.4161/cc.5.8.2687
Abstract | PDF

Loss of the G1/S checkpoint is recognized as a mandatory step in development of cancer. Nevertheless, both in vivo and in vitro experiments have indicated that this condition highly sensitizes cells to apoptosis. E.g., primary mouse embryonic fibroblasts that lack the complete retinoblastoma suppressor gene family (TKO MEFs) massively die under mitogen-deprived conditions. The prevailing hypothesis therefore is that the increased proliferative capacity of cells that have lost the G1/S checkpoint becomes apparent by suppression of apoptosis. However, this view was recently challenged by the finding that suppression of apoptotic cell death in TKO MEFs did not allow unconstrained proliferation; instead, cells became arrested in G2. This mechanism, which is dependent on p53, provides yet another barrier to oncogenic transformation. Thus, progression to malignancy of Rb-deficient lesions by alleviation of G2 arrest may offer an alternative explanation for the synergism between loss of Rb and p53 in tumorigenesis.

Open Access Article

microRNA Control of Lifespan and Metabolism

Michelle Boehm and Frank J. Slack
Pages 837 - 840
http://dx.doi.org/10.4161/cc.5.8.2688
Abstract | PDF

A family of small, non-coding RNAs, known as microRNAs, has recently emerged as sequence-specific regulators of gene expression. Hundreds of microRNAs have been identified in almost all metazoans genomes, but they are only beginning to be classified by functional roles. Here, we review microRNAs that have been shown to play roles in two closely related processes, lifespan and metabolic regulation. Understanding the metabolic and lifespan regulatory roles of these novel gene regulators will undoubtedly further our understanding of the complex genetic networks that control lifespan and metabolism, and will also provide us with novel targets for the theurapeutic intervention of metabolic and age-related diseases.

Open Access Article

Friend or Foe? The Proteasome in Combined Cancer Therapy

Dennis Sohn, Gudrun Totzke, Klaus Schulze-Osthoff and Reiner U. Jänicke
Pages 841 - 845
http://dx.doi.org/10.4161/cc.5.8.2740
Abstract | PDF

The proteasome is a multicatalytic enzyme complex that is responsible for degradation of the vast majority of intracellular proteins. Thus, it is involved in diverse cellular processes such as proliferation, differentiation and apoptosis. Especially its latter function yielded in the development of specific proteasomal inhibitors which have recently entered clinical trials due to their tremendous apoptosis-inducing capability. However, several recent studies including ours provided substantial evidence that a combined treatment of tumors with apoptosis-inducing agents and proteasomal inhibitors might even cause adverse effects leading to a prolonged survival of tumor cells. Based on our model of a biphasic role for the proteasome in apoptosis, we believe that a successful combat of tumors that relies on a combinational therapy with proteasomal inhibitors requires careful evaluation of several critical aspects in order to avoid a friend becoming a foe.




REVIEWS

Open Access Article

Regulatory Molecular Biology

Arthur B. Pardee
Pages 846 - 852
http://dx.doi.org/10.4161/cc.5.8.2634
Abstract | PDF

Close regulations of molecular biological processes are essential for life. Defective controls cause diseases such as cancer and neurological malfunctions. We now are provided with a plethora of regulatory mechanisms exerted at many levels. Prominent are covalent protein modifications, non-covalent feedback inhibition that modifies enzyme activity, and enzyme induction. Non-covalent or covalent binding to them of either small molecules or proteins act on functional DNA, RNA, proteins and metabolites regulates their production and degradation rates, activities and intra-cell locations. Time frames differ greatly, from seconds to days or longer. A control at every level is balanced by an opposing mechanism: populations of organisms are balanced by birth vs. death, cell synthesis by apoptosis, mutation by DNA repair, macromolecular syntheses by their degradations, metabolite anabolism vs. catabolism, enzyme activation by inhibition, protein kinases by phosphatases. Any abnormal molecular condition is sensed when regulation is defective as in cancer, which leads to its rectification, to cell death, or to disease if this is not possible.

Open Access Article

Polo-Like Kinases: A Team in Control of the Division

Barbara C.M. van de Weerdt and Rene H. Medema
Pages 853 - 864
http://dx.doi.org/10.4161/cc.5.8.2692
Abstract | PDF

Polo, the founding member of the family of polo-like kinases (Plks) was identified in a Drosphila screen for mutants affecting spindle pole behavior.1 Several mutants showed defects at their spindle poles and were hence named after the magnetic poles of the earth or geo-magnetic phenomena associated with them, like Polo and Aurora. Currently, the conserved family of Plks consists of many members throughout various species. Multiple Plks are present in mammalian cells (Plk1, Plk2/Snk, Plk3/Fnk/Prk, and Plk4/Sak) and Xenopus (Plx1-3), whereas in other species only one member has been identified, like Polo in Drosophila, Cdc5 in budding yeast and Plo1 in fission yeast. Plks are now viewed as important regulators of multiple functions before and during the mitotic cell division. In this review, we will focus our attention on human Plk1 and its family members Plk2-4 and the many roles they play during mitosis. Furthermore, we will describe the currently knowledge of the regulation of these functions.




BRIEF REPORTS

Open Access Article

The Fission Yeast APC Activator Ste9 is Regulated by mRNA Decay

Beatriz Álvarez, Miguel A. Blanco and Sergio Moreno
Pages 865 - 868
http://dx.doi.org/10.4161/cc.5.8.2647
Abstract | PDF

Fission yeast Ste9/Srw1 is a family member of the Fizzy-related APC activators that promote the ubiquitination and degradation of mitotic cyclins and other substrates at the end of mitosis and G1. These proteins are highly regulated during the cell cycle at the level of gene transcription and protein phosphorylation in order to guarantee the correct order of events during the cell cycle. Here we propose mRNA decay as a novel mechanism that regulates ste9+ gene expression during the cell cycle. We have characterized the elements in the 3´UTR of the ste9 mRNA responsible for this mechanism. Moreover, we demonstrate that the instability of ste9 mRNA is important for down-regulating Ste9 levels in G2, allowing appropriate cyclin B accumulation to promote timely entry into mitosis.

Open Access Article

A Late Origin of DNA Replication in the Trinucleotide Repeat Region of the Human FMR2 Gene

Paul D. Chastain II, Stephanie M. Cohen, Bruna P. Brylawski, Marila Cordeiro-Stone and David G. Kaufman
Pages 869 - 872
http://dx.doi.org/10.4161/cc.5.8.2648
Abstract | PDF

We confirmed that the replication of the fragile-X E site (FRAXE) in human chromosomal band Xq28 occurs at six hours into the eight-hour S phase of normal human fibroblasts. In this late-replicating region, we mapped an origin of DNA replication within the promoter of FMR2. This origin is coincident with CpG islands, the trinucleotide repeat, and exon 1 of the FMR2 gene. Identification of this origin may aid in the investigation of the mechanism of trinucleotide repeat expansion and its effect on FMR2 expression. In addition, knowledge of the chromosomal locations and sequence characteristics of both early and late origins of DNA replication, such as the one described in this report, will facilitate studies of the molecular determinants of the time of activation of different origins of replication and allow us to refine our insights concerning origin inactivation in response to the DNA damage-induced intra-S checkpoint.

Open Access Article

Control of AIF-mediated Cell Death by the Functional Interplay of SIRT1 and PARP-1 in Response to DNA Damage

Ullas Kolthur-Seetharam, Françoise Dantzer, Michael W. McBurney, Gilbert de Murcia1 and Paolo Sassone-Corsi
Pages 873 - 877
http://dx.doi.org/10.4161/cc.5.8.2690
Abstract | PDF

Cell survival after genotoxic stress is determined by a counterbalance of pro- and anti-death factors. Sirtuins (SIRTs) are deacetylases that promote cell survival whereas poly(ADP-ribose) polymerases (PARPs) can act both as survival and death inducing factor and the two protein families are strictly dependent on NAD+ for their activities. Here we report that SIRT1 modulates PARP-1 activity upon DNA damage. Activation of SIRT1 by resveratrol leads to reduced PARP-1 activity and there is a drastic increase in PAR synthesis in sirt1-null cells. The unbalanced regulation of PARP-1 in the absence of SIRT1 results in AIF (apoptosis inducing factor)-mediated cell death. Our findings establish a functional link between the two NAD+-dependent enzyme systems and provide a physiological interpretation for the mechanism of death in cells lacking SIRT1.




REPORTS

Open Access Article

2-Deoxy-D-glucose Reduces the Level of Constitutive Activation of ATM and Phosphorylation of Histone H2AX

Toshiki Tanaka, Akira Kurose, H. Dorota Halicka, Frank Traganos and Zbigniew Darzynkiewicz
Pages 878 - 882
http://dx.doi.org/10.4161/cc.5.8.2681
Abstract | PDF

Histone H2AX phosphorylated on Ser-139, defined as γH2AX, is a reporter of DNA double-strand breaks (DSBs). While H2AX undergoes phosphorylation after induction of DNA damage by genotoxic agents or during physiological events that involve DNA recombination, it also is phosphorylated in untreated normal and tumor cells. We recently reported that this constitutive H2AX phosphorylation (CHP) is markedly reduced by the antioxidant N-acetyl-L-cysteine (NAC), and postulated that it reflects the oxidative DNA damage (“endogenous DSBs”) induced by reactive oxygen species (ROS) generated by metabolic activity during progression through the cell cycle. In the present study, we provide evidence that growth of cells from three human lymphoblastoid cell lines TK6, NH32 and WTK1 in the presence of the glucose antimetabolite 2-deoxy-D-glucose (2-DG) led to a distinct reduction in the level of CHP. The reduction of CHP was more pronounced in S and G2M than in G1 phase cells. Constitutive activation of ATM was also reduced. The data suggest that a decrease in a cell’s metabolic activity as a result of inhibition of glycolysis by 2-DG reduces generation of ROS which leads to the reduction of oxidative DNA damage. The data also point out that ATM may play a role in CHP induced by oxidative DNA damage. Therefore, the assay of CHP by multiparameter cytometry provides the means to measure effects of antioxidants and metabolic inhibitors on endogenous oxidative DNA damage in relation to cell cycle phase.

Open Access Article

M-phase MELK activity is regulated by MPF and MAPK

Caroline Badouel, Roman Körner, Marie Frank-Vaillant, Anne Couturier, Erich A. Nigg and Jean-Pierre Tassan
Pages 883 - 889
http://dx.doi.org/10.4161/cc.5.8.2683
Abstract | PDF

Caroline Badouel, Roman Körner, Marie Frank-Vaillant, Anne Couturier, Erich A. Nigg and Jean-Pierre TassanThe protein kinase MELK is implicated in the control of cell proliferation, cell cycle and mRNA splicing. We previously showed that MELK activity is correlated with its phosphorylation level, is cell cycle dependent, and maximal during mitosis. Here we report on the identification of T414, T449, T451, T481 and S498 as residues phosphorylated in Xenopus MELK (xMELK) in M-phase egg extract. Phosphorylations of T449, T451, T481 are specifically detected during mitosis. Results obtained in vivo showed that MPF and MAPK pathways are involved in xMELK phosphorylation. In vitro, MPF and MAPK directly phosphorylate xMELK and MPF phosphorylates xMELK on T481. In addition, phosphorylation by MPF and MAPK enhances MELK activity in vitro. Taken together our results indicate that MELK phosphorylation by MPF and MAPK enhance its activity during M-phase.

Open Access Article

Posttranslational Regulation of Circadian Transcriptional Clock (NPAS2)/BMAL1 Complex by Cryptochromes

Roman V. Kondratov, Anna A. Kondratova, Choogon Lee, Victoria Y. Gorbacheva, Mikhail V. Chernov and Marina P. Antoch
Pages 890 - 895
http://dx.doi.org/10.4161/cc.5.8.2684
Abstract | PDF

Mammalian CLOCK(NPAS2), BMAL1 and CRYPTOCHROMEs are core components of the circadian oscillatory mechanism. The active CLOCK/BMAL1 or NPAS2/BMAL1 complexes regulate expression of numerous genes including two Cryptochromes. The products of these genes, CRY1 and CRY2, in turn repress CLOCK/BMAL1 transcriptional activity by an unknown mechanism. We have examined the effect of CRYPTOCHROMEs on posttranslational modifications and intracellular distribution of endogenous and ectopically expressed CLOCK(NPAS2) and BMAL1 proteins. We found that ectopic coexpression with CRY led to stabilization and nuclear accumulation of unphosphorylated forms of the proteins, which directly correlated with the inhibition of their transcriptional activity. This effect was CRY-specific, as other known repressors of CLOCK/BMAL1 and NPAS2/BMAL1 transcriptional activity were not able to induce similar effects. CRYs had no effect on CLOCK(NPAS2)/BMAL1 complex formation or its ability to bind DNA. Altogether, these results demonstrate that CRYs regulate the functional activity of circadian transcriptional complex at the posttranslational level. Importantly, the posttranslational modifications and intracellular distribution of CLOCK and BMAL1 proteins were critically impaired in the tissues of mice with targeted disruption of both Cry genes, thus confirming the suggested role of CRY in clock function in vivo. Based on these findings we propose a modified model of the circadian transcriptional control, which implies CRY-mediated periodic rotation of transcriptionally active and inactive forms of CLOCK/BMAL1 on the promoter. This model provides mechanistic explanation for previously reported dual functional activity of CLOCK/BMAL1 and highlights the involvement of the circadian system in modulating the organism’s response to various types of genotoxic stress, including chemotherapy and radiation.

Open Access Article

Human Sgo1 Is An Excellent Target For Induction of Apoptosis Of Transformed Cells

Yali Yang, Xiaoxing Wang and Wei Dai
Pages 896 - 901
http://dx.doi.org/10.4161/cc.5.8.2691
Abstract | PDF

Aneuploidy, a frequent form of genomic instability, is a direct result of mis-segregation of sister chromatids during mitosis. Cohesion of sister chromatids in eukaryotes is primarily achieved by the cohesin complex. Since Sgo1 functions to protect centromeric cohesin and since depletion of Sgo1 results in precocious chromosomal segregation and massive mitotic arrest, we reasoned that disruption of Sgo1 functions by delivery of a competitive peptide could interfere with normal cell cycle progression. Treatment of HeLa and A549 cells with a Sgo1 fragment of 22 amino acids (corresponding to a conserved basic region at the C-terminus) linked with a cell-permeable peptide penetratin, but not with penetratin alone, resulted in concentration-dependent suppression of viability. Following 24 h treatment with Sgo1 peptide at a 40 _M final concentration, HeLa cell viability was dramatically reduced. The Sgo1 peptide induced rapid and drastic changes of cell and nuclear morphologies, which were associated with disappearance of cellular Sgo1 at kinetochores in mitotic cells. Moreover, the reduction in HeLa cell viability after treatment with Sgo1 peptide resulted from induction of apoptosis. Interestingly, WI-38 diploid fibroblasts were rather resistant to the treatment with Sgo1 peptide. These normal cells exhibited little changes of cellular and nuclear morphologies, and remained viable after exposure to 40 _M of Sgo1 peptide for 24 h. Our combined studies thus suggest that small peptides corresponding to conserved regions of Sgo1 are capable of functioning as dominant negative inhibitors in vivo and that Sgo1 is an excellent target for induction of apoptosis of cancer cells.